Heroin addiction is a disease of chronic relapse that harms the individual through devaluation of personal duties and relationships in favor of finding and using drugs. Heroin in particular is disconcerting given its recent rise of use in the United States, and a new study labeling heroin the second most dangerous drug to the user and society [1]. Given the adverse consequences of heroin use, the goal to be drug free holds great importance for the user. Unfortunately, the incidence of relapse is quite high. Thus the question arises: Why do individuals have different relapse outcomes? A number of reports have shown a large interplay between environmental and genetic factors that contribute to the variability in relapse outcomes. Therefore the current proposal will examine how individual differences affect relapse outcome across multiple time periods in Specific Aim 1. Our laboratory has considerable experience in tracking individual differences in sensitivity to drugs of abuse by examining to what extent an animal devalues a drug paired natural reward cue. Rodents that greatly avoid a drug paired natural reward cue display increased motivation to work and take drug [2, 3]. This is the case for a heroin-paired cue as well. Epigenetic differences may contribute to such individual differences in response to drug [4], and DNA methylation, in particular, has been examined as potential factor for relapse given its perceived stability. Nevertheless, current DNA methylation studies have only scratched the surface. No studies, to our knowledge, have examined the DNA methylation differences in rats that respond differently to heroin or determined whether these changes are truly permanent during abstinence. Specific Aim 3, then, will use the rats from Specific Aim 1 to determine whether differential responding to heroin is linked to differences in DNA methylation in brain. Finally, the learning and memory field has shown that environmental enrichment can alter the epigenetic the landscape and our data [5], and those of others [6-9], have shown that enrichment can protect against addiction-like behaviors. Therefore, a next step is to determine if enrichment status can protect a rat from not only drug-induced reinstatement (Specific Aim 2), but also the accompanying epigenetic changes. Specific Aim 3 will use the brains from rats from Specific Aim 2 to address this question. In collaboration with the Freeman laboratory, we recently developed a novel next generation sequencing approach to loci-specific DNA methylation analysis and obtained pilot data linking increased responsiveness to heroin in the model with a distinct hippocampal DNA methylation prolife. From this gap in knowledge, I seek to understand how an individual's response to heroin affects abstinence and relapse and link these individual differences in vulnerability with DNA methylation.